Efficacy of personalized rTMS to enhance upper limb function in subacute stroke patients: a protocol for a multi-center, randomized controlled study.

Background: Repetitive transcranial magnetic stimulation (rTMS) is widely used therapy to enhance motor deficit in stroke patients. To date, rTMS protocols used in stroke patients are relatively unified. However, as the pathophysiology of stroke is diverse and individual functional deficits are distinctive, more precise application of rTMS is warranted. Therefore, the objective of this study was to determine the effects of personalized protocols of rTMS therapy based on the functional reserve of each stroke patient in subacute phase. Methods: This study will recruit 120 patients with stroke in subacute phase suffering from the upper extremity motor impairment, from five different hospitals in Korea. The participants will be allocated into three different study conditions based on the functional reserve of each participant, measured by the results of TMS-induced motor evoked potentials (MEPs), and brain MRI with diffusion tensor imaging (DTI) evaluations. The participants of the intervention-group in the three study conditions will receive different protocols of rTMS intervention, a total of 10 sessions for 2 weeks: high-frequency rTMS on ipsilesional primary motor cortex (M1), high-frequency rTMS on ipsilesional ventral premotor cortex, and high-frequency rTMS on contralesional M1. The participants of the control-group in all three study conditions will receive the same rTMS protocol: low-frequency rTMS on contralesional M1. For outcome measures, the following assessments will be performed at baseline (T0), during-intervention (T1), post-intervention (T2), and follow-up (T3) periods: Fugl-Meyer Assessment (FMA), Box-and-block test, Action Research Arm Test, Jebsen-Taylor hand function test, hand grip strength, Functional Ambulatory Category, fractional anisotropy measured by the DTI, and brain network connectivity obtained from MRI. The primary outcome will be the difference of upper limb function, as measured by FMA from T0 to T2. The secondary outcomes will be the differences of other assessments. Discussion: This study will determine the effects of applying different protocols of rTMS therapy based on the functional reserve of each patient. In addition, this methodology may prove to be more efficient than conventional rTMS protocols. Therefore, effective personalized application of rTMS to stroke patients can be achieved based on their severity, predicted mechanism of motor recovery, or functional reserves. Clinical trial registration: https://clinicaltrials.gov/, identifier NCT06270238.

An Exercise Prescription for Patients with Stroke and Sarcopenia Based on the Modified Delphi Study.

BACKGROUND: We aimed to develop a consensus on the need for and priorities of exercise to treat preexisting sarcopenia with hemiplegic stroke. METHODS: A modified three-round Delphi study was conducted. The panelists responded to the questionnaire on a 7-point Likert scale. Responses were returned with descriptive statistics in the next round. Consensus was defined as >75% agreement (score of 5-7) with a median > 5. The percentage of strong agreement (score of 6-7) and Kendall's coefficient of concordance were calculated to demonstrate a more refined interpretation of the consensus. RESULTS: Fifteen panelists contributed to all rounds. The need for exercise was demonstrated. The consensus was reached on 53 of 58 items in the first round and all items in the second and final rounds. The percentage of strong agreement was high for all but eight items. CONCLUSIONS: This study is the first Delphi study to investigate the need for and priorities of exercise for treating preexisting sarcopenia in stroke hemiplegia. We present a standard recommendation including 57 priorities and a strong recommendation including 49 priorities. The eight items that were excluded reflected factors that are less important to hemiplegic patients with poor balance, cognitive decline, or mental vulnerability.

Profiling the Phytochemicals of Orostachys margaritifolia: Biological Activities, LC-ESI/MS, and HPLC Analyses.

Orostachys margaritifolia Y. N. Lee (OMY) is an endemic Korean plant in the family Crassulaceae that is known to contain a variety of bioactive compounds. To assess the physiological activities of an OMY ethanol extract, ABTS+ and DPPH radical scavenging assays and a nitric oxide (NO) inhibition assay were conducted. The phytochemical makeup of the extract was profiled via liquid chromatography-mass spectrometry (LC-ESI/MS) and high-performance liquid chromatography with a photodiode array detector (HPLC/PDA). The OMY extract was found to have weaker ABTS+ and DPPH radical scavenging activities than the control group (green tea). In the NO inhibition assay, the OMY extract induced a significant increase in macrophage cell viability but showed a lower NO inhibitory activity than l-NAME, producing an IC50 value of 202.6 µg/mL. The LC-ESI/MS and HPLC/PDA analyses identified isoquercitrin and astragalin in the OMY extract, quantifying their contents at 3.74 mg/g and 3.19 mg/g, respectively. The study revealed possibilities for the utilization of OMY as a future source of drugs for alleviating inflammation and diseases related to reactive oxygen species.

Electric field simulation and appropriate electrode positioning for optimized transcranial direct current stimulation of stroke patients: an in Silico model.

Transcranial Direct Current Stimulation (tDCS) has benefits for motor rehabilitation in stroke patients, but its clinical application is limited due to inter-individual heterogeneous effects. Recently, optimized tDCS that considers individual brain structure has been proposed, but the utility thereof has not been studied in detail. We explored whether optimized tDCS provides unique electrode positions for each patient and creates a higher target electric field than the conventional approach. A comparative within-subject simulation study was conducted using data collected for a randomized controlled study evaluating the effect of optimized tDCS on upper extremity function in stroke patients. Using Neurophet tES LAB 3.0 software, individual brain models were created based on magnetic resonance images and tDCS simulations were performed for each of the conventional and optimized configurations. A comparison of electrode positions between conventional tDCS and optimized tDCS was quantified by calculation of Euclidean distances. A total of 21 stroke patients were studied. Optimized tDCS produced a higher electric field in the hand motor region than conventional tDCS, with an average improvement of 20% and a maximum of 52%. The electrode montage for optimized tDCS was unique to each patient and exhibited various configurations that differed from electrode placement of conventional tDCS. Optimized tDCS afforded a higher electric field in the target of a stroke patient compared to conventional tDCS, which was made possible by appropriately positioning the electrodes. Our findings may encourage further trials on optimized tDCS for motor rehabilitation after stroke.

Safety and therapeutic effects of personalized transcranial direct current stimulation based on electrical field simulation for prolonged disorders of consciousness: study protocol for a multi-center, double-blind, randomized controlled trial.

Background: Disorders of consciousness (DOC) resulting from acquired brain injury (ABI) increase the mortality rate of patients, complicate rehabilitation, and increase the physical and economic burden that DOC imposes on patients and their families. Thus, treatment to promote early awakening from DOC is vital. Transcranial direct current stimulation (tDCS) has shown great potential for promoting neuro-electrochemical activity. However, previous tDCS studies did not consider structural damage or head and brain lesions, so the applicability of the results to all DOC patients was limited. In this study, to establish a patient-specific tDCS treatment plan considering the brain lesions of and damage sustained by DOC patients, we considered the electric field calculated by a the "finite electric" three-dimensional brain model based on magnetic resonance images. This protocol was developed to aid tDCS treatment of actual patients, and to verify its safety and effectiveness. Methods/design: Twenty-four patients with DOC after ABI will be enrolled in this cross-over trial. All participants will receive typical rehabilitation combined with sham tDCS and typical rehabilitation plus personalized tDCS (P-tDCS). Each interventional period will last 2 weeks (30 min/day, 5 days/week). The primary outcome [score on the Korean version of the Coma Recovery Scale-Revised (K-CRS-R)] will be assessed at baseline and the end of the first day of the intervention. Secondary outcomes (K-CRS-R at 1 week and 2 weeks after experimental session and quantitative EEG changes quantitative electroencephalography changes) will be measured at baseline and the end of week 4. Adverse events will be recorded during each treatment session. Discussion: For patients with neurological disorders, tDCS has served as a painless, non-invasive, easily applied, and effective therapy for several decades, and there is some evidence that it can improve the level of consciousness of patients with DOC. However, variability in the effects on consciousness among subjects have been reported and personalized strategies are lacking. This protocol is for a randomized controlled trial designed to validate the effectiveness and safety of P-tDCS combined with typical rehabilitation for DOC. Clinical trial registration: https://cris.nih.go.kr, identifier KCT0007157.

Structural Integrity of the Cerebellar Outflow Tract Predicts Long-Term Motor Function After Middle Cerebral Artery Ischemic Stroke.

BACKGROUND: The cerebellum plays a crucial role in functional movement by influencing sensorimotor coordination and learning. However, the effects of cortico-cerebellar connectivity on the recovery of upper extremity motor function after stroke have not been investigated. We hypothesized that the integrity of the cortico-cerebellar connections would be reduced in patients with a subacute middle cerebral artery (MCA) stroke, and that this reduction may help to predict chronic upper extremity motor function. METHODS: We retrospectively analyzed the diffusion-tensor imaging of 25 patients with a subacute MCA stroke (mean age: 62.2 ± 2.7 years; 14 females) and 25 age- and sex-matched healthy controls. We evaluated the microstructural integrity of the corticospinal tract (CST), dentatothalamocortical tract (DTCT), and corticopontocerebellar tract (CPCT). Furthermore, we created linear regression models to predict chronic upper extremity motor function based on the structural integrity of each tract. RESULTS: In stroke patients, the affected DTCT and CST showed significantly impaired structural integrity compared to unaffected tracts and the tracts in controls. When all models were compared, the model that used the fractional anisotropy (FA) asymmetry indices of CST and DTCT as independent variables best predicted chronic upper extremity motor function (R2 = .506, P = .001). The extent of structural integrity of the CPCT did not significantly differ between hemispheres or groups and was not predictive of motor function. CONCLUSIONS: We found evidence that microstructural integrity of the DTCT in the subacute phase of an MCA stroke helped to predict chronic upper extremity motor function, independent of CST status.

A case of vocal cord dysfunction diagnosed in a 10-year-old girl with recurrent wheezing and dyspnea

Vocal cord dysfunction is one of the causes of dyspnea and is characterized by paradoxical closure of the vocal cords. The paradoxical movement of the vocal cords produces the limitation of airflow, resulting dyspnea, chest tightening, hoarseness, stridor, or wheezing. These findings are similar to those of other upper airway obstruction diseases or asthma; therefore, a high index of suspicion and clear differential diagnosis are required. Here, we discuss a case of vocal cord dysfunction aged 10 years that presented recurrent wheezing and dyspnea. The abnormal movement of the vocal cords was observed by fiberoptic laryngotracheobronchoscopy, which was correlated with stridor during respiration. Repeated episodic symptoms were controlled by the multidisciplinary team approach; however, surgical treatment was needed to stabilize the symptom.

MRI-Based Personalized Transcranial Direct Current Stimulation to Enhance the Upper Limb Function in Patients with Stroke: Study Protocol for a Double-Blind Randomized Controlled Trial.

Transcranial direct current stimulation (tDCS) has been shown to have the potential to improve the motor recovery of the affected upper limbs in patients with stroke, and recently, several optimized tDCS methods have been proposed to magnify its effectiveness. This study aims to determine the effectiveness of personalized tDCS using brain MRI-based electrical field simulation and optimization, to enhance motor recovery of the upper limbs in the patients. This trial is a double-blind, randomized controlled trial in the subacute to chronic rehabilitation phase. Forty-two adult stroke patients with unilateral upper limb involvement will be randomly allocated to three groups: (1) personalized tDCS with MRI-based electrical field simulation and optimized stimulation, (2) conventional tDCS with bihemispheric stimulation of the primary motor cortex, and (3) sham tDCS. All three groups will undergo 10 intervention sessions with 30 min of 2-mA intensity stimulation, during a regular upper limb rehabilitation program over two weeks. The primary outcome measure for the motor recovery of the upper limb impairment is the Fugl-Meyer assessment for the upper extremity score at the end of the intervention, and the secondary measures include changes in the motor evoked potentials, the frequency power and coherence of the electroencephalography, performance in activities of daily living, and adverse events with a 1-month follow-up assessment. The primary outcome will be analyzed on the intention-to-treat principle. There is a paucity of studies regarding the effectiveness of personalized and optimized tDCS that considers individual brain lesions and electrical field characteristics in the real world. No feasibility or pivotal studies have been performed in stroke patients using brain MRI, to determine a lesion-specific tDCS simulation and optimization that considers obstacles in the segmentation and analysis of the affected brain tissue, such as ischemic and hemorrhagic lesions. This trial will contribute to addressing the effectiveness and safety of personalized tDCS, using brain MRI-based electrical field simulation and optimization, to enhance the motor recovery of the upper limbs in patients with stroke.

Physical activity, quality of life and parenting stress in children with cerebral palsy.

BACKGROUND: Children with cerebral palsy (CP) are less physically active than their typically developing peers. The effects of decreased physical activity on children's quality of life (QOL) or caregiver's lives are not well understood. This study aimed to investigate the association between physical activity, QOL, and parenting stress in children with CP. METHODS: A prospective cross-sectional study was done in children with CP. Daily physical activity was measured over 7 days using an accelerometer (ActiGraph™). Caregivers completed the Child Health Questionnaire - Parent Form 50 and Parenting Stress Index - Short Form. Multiple regression analysis was used. RESULTS: In total, data from 65 children with Gross Motor Function Classification System (GMFCS) levels I-V, aged 4-13 years, were analyzed. Non-ambulatory (GMFCS IV-V) children had significantly lower activity counts and moderate to vigorous physical activity (MVPA) in comparison with ambulatory (GMFCS I-II) and marginally ambulatory (GMFCS III) children. The ambulatory group had better physical QOL than the non-ambulatory group. Caregivers of the non-ambulatory or marginally ambulatory children with CP reported higher parenting stress levels than those of the ambulatory group. Time spent in MVPA and activity counts were positively associated with physical QOL in children with CP. Low activity counts and low amounts of MVPA of children with CP were significant predictors of high parenting stress. CONCLUSIONS: Physical activity in children with CP is associated with children's QOL and parenting stress. These results suggest that clinicians should conduct physical activity assessment and counseling to promote physical activity.

Clinical Course of Dysphagia in Patients with Nemaline Myopathy.

Nemaline myopathy (NM) is a rare congenital myopathy, a group of disorders that are clinically and genetically heterogeneous. Infants and children with NM often suffer from recurrent pulmonary infections and swallowing difficulty, leading to malnutrition. However, knowledge about the clinical course and prognosis of dysphagia is limited. In this study, we reported the clinical course of two NM patients suffering from dysphagia. Although tube feeding was required for several months after birth, it was eventually possible to obtain sufficient nutrition with an oral diet. Therefore, dysphagia rehabilitation therapy through a series of evaluations should be considered even in children with severe oral motor dysfunction. Through these cases, physicians should be convinced that the symptoms of dysphagia in children with NM can be improved and be able to encourage their parents by explaining this progress. They have the potential to show improvements in swallowing function and will finally be able to take food slowly but fully orally.

Preservation of Cerebellar Afferent Pathway May Be Related to Good Hand Function in Patients with Stroke.

Many chronic stroke patients suffer from worsened hand function, and functional recovery of the hand does not occur well after six months of stroke. Therefore, predicting final hand function after stroke through acute phase imaging would be an important issue in counseling with the patients or their family. Thus, we investigated the remaining white matter integrity in the corticospinal tract (CST) and cortico-ponto-cerebellar tract (CPCT) at the acute stage of stroke and chronic hand function after stroke, and present the cut-off value of fiber number (FN) and fractional anisotropy (FA) of CST and CPCT at the acute stage for predicting final hand function after the recovery period. This retrospective case-control study included 18 stroke patients who were classified into two groups: poor hand function with stroke (n = 11) and good hand function with stroke (n = 7). DTI was done within two months ± 15 days after onset, and the Jebson's Hand Function test was conducted 6-12 months after onset. The investigation of white matter was focused on the values of FN and FA for CST and CPCT, which were measured separately. The normalized (affected/non-affected) FA and FN values in the CPCT in the good hand function group were higher than those in the poor hand function group. The normalized FN and FA values in the CST were not significantly different between the poor hand function group and the good hand function group. The normalized cut-off value that distinguished the good hand function group from the poor hand function group was 0.8889 for FA in the CPCT. The integrity of the CPCT in the acute stage was associated with hand function in the chronic stage after a stroke. Ultimately, the integrity of the CPCT in the early stage after onset can be used to predict chronic hand function. Based on these results, cerebellar afferent fiber measurements may be a useful addition to predict hand function and plan specific rehabilitation strategies in stroke patients.

Factors Influencing the Progression and Direction of Scoliosis in Children with Neurological Disorders.

(1) Background: scoliosis is highly prevalent in children with neurological disorders, however, studies predicting the progression and affecting the direction of scoliosis have been insufficient. We investigated the factors associated with the progression and direction of scoliosis in children with neurological disorders. (2) Method: retrospectively, 518 whole spine radiographs from 116 patients were used for analysis. Factors affecting the progression of scoliosis over time were analyzed using linear mixed-effects model. Factors associated with the apex direction of the scoliosis were analyzed. (3) Results: pelvic obliquity (PO) ≥ 2.5°, gross motor function classification system level V, vertebral rotation, and female sex significantly affect the progression of scoliosis (p = 0.04, <0.001, <0.001, 0.005, respectively). The higher side of PO and the apex side of scoliosis were interrelated (χ² = 14.58, p < 0.001), but the asymmetrical neurological upper extremity involvement was not. (4) Conclusions: severely impaired gross motor function, PO, vertebral rotation, and female sex were significantly related to the progression of scoliosis. The higher side of PO was opposite to the apex side of scoliosis. By identifying the factors that influence the progression of scoliosis, patients at high risk could be more actively intervened to minimize the severe complications.

Alteration of White Matter in Patients with Central Post-Stroke Pain.

A stroke may be followed by central post-stroke pain (CPSP), which is characterized by chronic neuropathic pain. The exact mechanism has not yet been fully uncovered. We investigated alterations in the white matters in patients with CPSP, compared with stroke patients without CPSP and normal controls. Our retrospective cross-sectional, case-control study participants were assigned to three groups: CPSP (stroke patients with CPSP (n = 17)); stroke control (stroke patients without CPSP (n = 26)); and normal control (normal subjects (n = 34)). The investigation of white matter for CPSP was focused on the values of fiber numbers (FN) and fractional anisotrophy (FA) for spinothalamic tract (STT), anterior thalamic radiation (ATR), superior thalamic radiation (STR) and posterior thalamic radiation (PTR), and corticospinal tract (CST) was measured. The FA for the STT and STR of the CPSP group were lower than those for the stroke control and normal control groups. The FA of CST and ATR did not differ between the CPSP and stroke groups, but both differed from the normal control. The FA of PTR in the stroke control group differed from the normal control group, but not from the CPSP group. The FN of CST, STT, ATR, and STR for the CPSP and stroke control groups did not differ from each other, but both differed from those of normal controls. FN of PTR did not differ between the CPSP and normal control groups. The alterations in the spinothalamic tract and superior thalamic radiation after stroke would play a role in the pathogenesis of CPSP.

The Long-Term Functional Effect of Thrombectomy on Patients with Middle Cerebral Artery Occlusion Who Exhibit Moderate to Severe Disability.

Background and Objectives: Endovascular thrombectomy (EVT is an emerging gold standard treatment for acute cerebral infarction and may allow functional improvement after subacute cerebral infarction. However, the long-term functional benefits of EVT in patients with moderate to severe disability remain unclear. We investigated the effects of EVT on the activities of daily living (ADL), handicap, gait, and eating in patients with middle cerebral artery (MCA) occlusion who exhibited moderate to severe disability (score of 3-5 on the modified Rankin scale (mRS)) due to stroke, up to six months after onset. Materials and Methods: This retrospective longitudinal case-control study assessed 45 patients with MCA occlusion who exhibited moderate to severe disability (mRS score ≥ 3): 15 underwent EVT and 30 served as controls. Clinical assessments were conducted at two weeks (12-16 days), four weeks (26-30 days), and six months (180-210 days) after stroke onset. Functional assessments comprised the Korean version of the modified Barthel index (MBI), mRS, functional ambulation category (FAC), and dysphagia outcome severity scale (DOSS) to assess disability, handicap, gait, and eating. Results: The MBI, mRS, FAC, and DOSS scores all improved significantly (all p < 0.05) in the EVT group, compared to the controls. Conclusions: EVT has favorable effects on performing routine ADL, the handicap itself, walking, and eating. Therefore, EVT is recommended for patients with acute MCA occlusion, including those with severe disability at the initial assessment.

Is Lateropulsion Really Related with a Specific Lesion of the Brain?

Lateropulsion (pusher syndrome) is an important barrier to standing and gait after stroke. Although several studies have attempted to elucidate the relationship between brain lesions and lateropulsion, the effects of specific brain lesions on the development of lateropulsion remain unclear. Thus, the present study investigated the effects of stroke lesion location and size on lateropulsion in right hemisphere stroke patients. The present retrospective cross-sectional observational study assessed 50 right hemisphere stroke patients. Lateropulsion was diagnosed and evaluated using the Scale for Contraversive Pushing (SCP). Voxel-based lesion symptom mapping (VLSM) analysis with 3T-MRI was used to identify the culprit lesion for SCP. We also performed VLSM controlling for lesion volume as a nuisance covariate, in a multivariate model that also controlled for other factors contributing to pusher behavior. VLSM, combined with statistical non-parametric mapping (SnPM), identified the specific region with SCP. Lesion size was associated with lateropulsion. The precentral gyrus, postcentral gyrus, inferior frontal gyrus, insula and subgyral parietal lobe of the right hemisphere seemed to be associated with the lateropulsion; however, after adjusting for lesion volume as a nuisance covariate, no lesion areas were associated with the SCP scores. The size of the right hemisphere lesion was the only factor most strongly associated with lateropulsion in patients with stroke. These results may be useful for planning rehabilitation strategies of restoring vertical posture and understanding the pathophysiology of lateropulsion in stroke patients.

Effect of Fascia Penetration in Lateral Femoral Cutaneous Nerve Conduction.

OBJECTIVE: To evaluate the effect of fascia penetration and develop a new technique for lateral femoral cutaneous nerve (LFCN) conduction studies based on the fascia penetration point (PP) identified using ultrasound. METHODS: The fascia PP of the LFCN was localized in 20 healthy subjects, and sensory nerve action potentials (SNAPs) were obtained at four different stimulation points-2 cm proximal to the PP (2PPP), PP, 2 cm distal to the PP (2DPP), and 4 cm distal to the PP (4DPP). We compared the stimulation technique based on the fascia penetration point (STBFP) with the conventional technique. RESULTS: The SNAP amplitude of the LFCN was significantly higher when stimulation was performed at the PP and 2DPP than at other stimulation points. Using the STBFP, SNAP responses were elicited in 38 of 40 legs, whereas they were elicited in 32 of 40 legs using the conventional technique (p=0.041). STBFP had a comparable SNAP amplitude and slightly delayed negative peak latency compared to the conventional technique. In terms of the time required, the time spent on STBFP showed a more consistent distribution than the time spent on the conventional technique (two-sample Kolmogorov-Smirnov test, p<0.05). CONCLUSION: SNAP of the LFCN significantly changed near the fascia PP, and stimulation at PP and at 2DPP provided high amplitudes. STBFP can help increase the response rate and ensure stable and consistent procedure time of the LFCN conduction study.

Multidisciplinary Approach to Decrease In-Hospital Delay for Stroke Thrombolysis.

BACKGROUND AND PURPOSE: Decreasing the time delay for thrombolysis, including intravenous thrombolysis (IVT) with tissue plasminogen activator and intra-arterial thrombectomy (IAT), is critical for decreasing the morbidity and mortality of patients experiencing acute stroke. We aimed to decrease the in-hospital delay for both IVT and IAT through a multidisciplinary approach that is feasible 24 h/day. METHODS: We implemented the Stroke Alert Team (SAT) on May 2, 2016, which introduced hospital-initiated ambulance prenotification and reorganized in-hospital processes. We compared the patient characteristics, time for each step of the evaluation and thrombolysis, thrombolysis rate, and post-thrombolysis intracranial hemorrhage from January 2014 to August 2016. RESULTS: A total of 245 patients received thrombolysis (198 before SAT; 47 after SAT). The median door-to-CT, door-to-MRI, and door-to-laboratory times decreased to 13 min, 37.5 min, and 8 min, respectively, after SAT implementation (P<0.001). The median door-to-IVT time decreased from 46 min (interquartile range [IQR] 36-57 min) to 20.5 min (IQR 15.8-32.5 min; P<0.001). The median door-to-IAT time decreased from 156 min (IQR 124.5-212.5 min) to 86.5 min (IQR 67.5-102.3 min; P<0.001). The thrombolysis rate increased from 9.8% (198/2,012) to 15.8% (47/297; P=0.002), and the post-thrombolysis radiological intracranial hemorrhage rate decreased from 12.6% (25/198) to 2.1% (1/47; P=0.035). CONCLUSIONS: SAT significantly decreased the in-hospital delay for thrombolysis, increased thrombolysis rate, and decreased post-thrombolysis intracranial hemorrhage. Time benefits of SAT were observed for both IVT and IAT and during office hours and after-hours.

Mono-uridylation of pre-microRNA as a key step in the biogenesis of group II let-7 microRNAs.

RNase III Drosha initiates microRNA (miRNA) maturation by cleaving a primary miRNA transcript and releasing a pre-miRNA with a 2 nt 3' overhang. Dicer recognizes the 2 nt 3' overhang structure to selectively process pre-miRNAs. Here, we find that, unlike prototypic pre-miRNAs (group I), group II pre-miRNAs acquire a shorter (1 nt) 3' overhang from Drosha processing and therefore require a 3'-end mono-uridylation for Dicer processing. The majority of let-7 and miR-105 belong to group II. We identify TUT7/ZCCHC6, TUT4/ZCCHC11, and TUT2/PAPD4/GLD2 as the terminal uridylyl transferases responsible for pre-miRNA mono-uridylation. The TUTs act specifically on dsRNAs with a 1 nt 3' overhang, thereby creating a 2 nt 3' overhang. Depletion of TUTs reduces let-7 levels and disrupts let-7 function. Although the let-7 suppressor, Lin28, induces inhibitory oligo-uridylation in embryonic stem cells, mono-uridylation occurs in somatic cells lacking Lin28 to promote let-7 biogenesis. Our study reveals functional duality of uridylation and introduces TUT7/4/2 as components of the miRNA biogenesis pathway.

TUT4 in concert with Lin28 suppresses microRNA biogenesis through pre-microRNA uridylation.

As key regulators in cellular functions, microRNAs (miRNAs) themselves need to be tightly controlled. Lin28, a pluripotency factor, was reported to downregulate let-7 miRNA by inducing uridylation of let-7 precursor (pre-let-7). But the enzyme responsible for the uridylation remained unknown. Here we identify a noncanonical poly (A) polymerase, TUTase4 (TUT4), as the uridylyl transferase for pre-let-7. Lin28 recruits TUT4 to pre-let-7 by recognizing a tetra-nucleotide sequence motif (GGAG) in the terminal loop. TUT4 in turn adds an oligouridine tail to the pre-let-7, which blocks Dicer processing. Other miRNAs with the same sequence motif (miR-107, -143, and -200c) are regulated through the same mechanism. Knockdown of TUT4 and Lin28 reduces the level of stem cell markers, suggesting that they are required for stem cell maintenance. This study uncovers the role of TUT4 and Lin28 as specific suppressors of miRNA biogenesis, which has implications for stem cell research and cancer biology.